Valeriana jatamansi Jones Inhibits Rotavirus-Induced Diarrhea via Phosphatidylinositol 3-Kinase/Protein Kinase B Signaling Pathway.

Rotavirus (RV), as the main cause of diarrhea in children under 5 years, contributes to various childhood diseases. Valeriana jatamansi Jones is a traditional Chinese herb and possesses antiviral effects. In this study we investigated the potential mechanisms of V. jatamansi Jones in RV-induced diarrhea. MTT assay was performed to evaluate cell proliferation and the diarrhea mice model was constructed using SA11 infection. Mice were administered V. jatamansi Jones and ribavirin. Diarrhea score was used to evaluate the treatment effect. The enzyme-linked immunosorbent assay was performed to detect the level of cytokines. Western blot and quantitative reverse transcription-PCR were used to determine protein and mRNA levels, respectively. Hematoxylin-eosin staining was applied to detect the pathological change of the small intestine. TdT-mediated dUTP nick-end labeling was conducted to determine the apoptosis rate. The results showed V. jatamansi Jones promoted MA104 proliferation. V. jatamansi Jones downregulated phosphatidylinositol 3-kinase (PI3K) and protein kinase B (AKT) in protein level, which was consistent with the immunohistochemistry results. Moreover, V. jatamansi Jones combined with ribavirin regulated interleukin-1ß (IL-1ß), interferon γ, IL-6, tumor necrosis factor α, and IL-10, and suppressed secretory immunoglobulin A secretion to remove viruses and inhibit dehydration. V. jatamansi Jones + ribavirin facilitated the apoptosis of small intestine cells. In conclusion, V. jatamansi Jones may inhibit RV-induced diarrhea through PI3K/AKT signaling pathway, and could therefore be a potential therapy for diarrhea.

Antiviral activities of resveratrol against rotavirus in vitro and in vivo.

BACKGROUND: Rotavirus (RV) is the primary causative agent for viral gastroenteritis among infants and young children worldwide. Currently, no clinically approved and effective antiviral drug for the treatment of RV infection is available. PURPOSE: We investigated the potential anti-RV activity of resveratrol and underlying mechanisms by which resveratrol acted against RV. METHODS: The anti-RV activity of resveratrol in vitro was evaluated using plaque reduction assays. The effects of resveratrol on yield of virion progeny, viral polyprotein expression and genomic RNA synthesis were respectively investigated using enzyme-linked immunosorbent assays, western blotting and qRT-PCR assays. Further, we also measured the antiviral effect of resveratrol by evaluation of antigen clearance and assessment of changes in proinflammatory cytokines/chemokines in RV-infected neonatal mouse model. RESULTS: Our results indicated that 20 µM of resveratrol significantly inhibited RV replication in Caco-2 cell line by suppressing RV RNA synthesis, protein expression, viroplasm plaque formation, progeny virion production, and RV-induced cytopathy independent of the different strains and cell lines of RV that we used. Analysis of the effect of time post-addition of resveratrol indicated that its application inhibited early processes in the RV replication cycle. Further study of the underlying mechanism of anti-RV activity indicated that resveratrol inhibited RV replication by suppressing expression of heat-shock protein 90 (HSP90) mRNA and protein, and that the effect occurred in a dose-dependent manner. Overexpression of HSP90 was found to have attenuated the inhibitory effect of resveratrol on RV replication. Interestingly, the application of resveratrol were found to down-regulate the level of inhibition of RV-mediated MEK1/2 and ERK phosphorylation. Using a RV-infected suckling mice model, we found that application of resveratrol significantly lessened the severity of diarrhea, decreased viral titers, and relieved associated symptoms. Levels of mRNA expression of interleukin-2, interleukin-10, tumor necrosis factor-α, interferon-γ, macrophage inflammatory protein 1, and monocyte chemotactic protein-1 were all found to have been sharply reduced in intestinal tissue from mice which had been treated with resveratrol (10 or 20 mg/kg) after RV infection (p < 0.05). CONCLUSION: These findings implied that resveratrol exhibits antiviral activity and could be a promising treatment for rotavirus infection.

Drug screening identifies gemcitabine inhibiting rotavirus through alteration of pyrimidine nucleotide synthesis pathway.

Although rotavirus infection is usually acute and self-limiting, it can cause chronic infection with severe diseases in immunocompromised patients, including organ transplantation recipients and cancer patients irrespective of pediatric or adult patients. Since no approved medication against rotavirus infection is available, this study screened a library of safe-in-man broad-spectrum antivirals. We identified gemcitabine, a widely used anti-cancer drug, as a potent inhibitor of rotavirus infection. We confirmed this effect in 2D cell cultures and 3D cultured human intestinal organoids with both laboratory-adapted rotavirus strains and five clinical isolates. Supplementation of UTP or uridine largely abolished the anti-rotavirus activity of gemcitabine, suggesting its function through inhibition of pyrimidine biosynthesis pathway. Our results support repositioning of gemcitabine for treating rotavirus infection, especially for infected cancer patients.

Screening for Components/Compounds with Anti-Rotavirus Activity: Detection of Interaction Between Viral Spike Proteins and Glycans.

Rotaviruses are the major etiologic agents of acute gastroenteritis. Viral attachment to the cell surface is crucial to initiate infection. The VP8∗ domain, the trypsinized cleavage fragment of the outermost spike protein VP4 of rotavirus, has a galectin-like structure required for binding to the cell surface. We used the evanescent-field fluorescence-assisted assay to understand the complex mechanism underlying the virus-glycan/glycoprotein interaction. Besides, we have described virus infection assays, neutralization assay, and pretreatment assay, using cell culture. These approaches using rotavirus particles will provide novel information that has been difficult to obtain from glycan microarray using recombinant VP8∗.

Vitamin D Alleviates Rotavirus Infection through a Microrna-155-5p Mediated Regulation of the TBK1/IRF3 Signaling Pathway In Vivo and In Vitro.

(1) Background: Vitamin D (VD) plays a vital role in anti-viral innate immunity. However, the role of VD in anti-rotavirus and its mechanism is still unclear. The present study was performed to investigate whether VD alleviates rotavirus (RV) infection through a microRNA-155-5p (miR-155-5p)-mediated regulation of TANK-binding kinase 1 (TBK1)/interferon regulatory factors 3 (IRF3) signaling pathway in vivo and in vitro. (2) Methods: The efficacy of VD treatment was evaluated in DLY pig and IPEC-J2. Dual-luciferase reporter activity assay was performed to verify the role of miR-155-5p in 1α,25-dihydroxy-VD3 (1,25D3) mediating the regulation of the TBK1/IRF3 signaling pathway. (3) Results: A 5000 IU·kg-1 dietary VD3 supplementation attenuated RV-induced the decrease of the villus height and crypt depth (p < 0.05), and up-regulated TBK1, IRF3, and IFN-ß mRNA expressions in the jejunum (p < 0.05). Incubation with 1,25D3 significantly decreased the RV mRNA expression and the RV antigen concentration, and increased the TBK1 mRNA and protein levels, and the phosphoprotein IRF3 (p-IRF3) level (p < 0.05). The expression of miR-155-5p was up-regulated in response to an RV infection in vivo and in vitro (p < 0.05). 1,25D3 significantly repressed the up-regulation of miR-155-5p in vivo and in vitro (p < 0.05). Overexpression of miR-155-5p remarkably suppressed the mRNA and protein levels of TBK1 and p-IRF3 (p < 0.01), while the inhibition of miR-155-5p had an opposite effect. Luciferase activity assays confirmed that miR-155-5p regulated RV replication by directly targeting TBK1, and miR-155-5p suppressed the TBK1 protein level (p < 0.01). (4) Conclusions: These results indicate that miR-155-5p is involved in 1,25D3 mediating the regulation of the TBK1/IRF3 signaling pathway by directly targeting TBK1.

Ursolic acid: A novel antiviral compound inhibiting rotavirus infection in vitro.

Rotavirus is one of the leading causes of severe acute gastroenteritis in children under 5 years of age, mainly affecting developing countries. Once the disease is acquired, no specific treatment is available; as such, the development of new drugs for effective antirotaviral treatment is critical. Ursolic acid is a pentacyclic triterpenoid with antiviral activity, which has been studied extensively in vitro and in vivo. To study the potential antirotaviral activity of ursolic acid, its toxic potential for viral particles (virucidal effect) and cultured cells (cytotoxicity) was analysed. No effect on virion infectivity was observed with treatments of up to 40 µM ursolic acid, while incipient cytotoxicity started to be evident with 20 µM ursolic acid. The antiviral potential of ursolic acid was evaluated in in-vitro rotavirus infections, demonstrating that 10 µM ursolic acid inhibits rotavirus replication (observed by a decrease in viral titre and the level of the main viral proteins) and affects viral particle maturation (a process associated with the endoplasmic reticulum) 15 h post infection. Interestingly, ursolic acid was also found to hamper the early stages of the viral replication cycle, as a significant reduction in the number and size of viroplasms, consistent with a decrease in VP6 and NSP2 viral proteins, was observed 4 h post infection. As such, these observations demonstrate that ursolic acid exhibits antiviral activity, suggesting that this chemical could be used as a new treatment for rotavirus.

Antiviral oxysterols are present in human milk at diverse stages of lactation.

Oxysterols are cholesterol oxidation derivatives. Those containing an additional hydroxyl group on the side chain of the cholesterol molecule result from a physiological enzymatic synthesis and include the majority of oxysterols present in the circulation. Among these, 25-hydroxycholesterol (25OHC) and 27-hydroxycholesterol (27OHC) are characterized by a broad antiviral activity and are now considered involved in the innate immune response against viruses. Despite the emerging role of these sterols in the innate antiviral defences, no data are available on their presence in human breast milk (BM) to date. In this study, we investigated the content of oxysterols of enzymatic synthesis in BM of twelve donor mothers at different stages of lactation (i.e. in colostrum, transitional milk, and mature milk) by gas chromatography-mass spectrometry analysis. The side-chain oxysterols 25OHC, 27OHC, and 24S-hydroxycholesterol (24SOHC) were actually present in BM in all stages of lactation, but the concentration of 27OHC showed a remarkable peak in colostrum. Antiviral assays revealed that all the colostrum samples contained 27OHC concentrations that were active in vitro against two relevant pediatric viral pathogens: the human rotavirus and the human rhinovirus. Overall, this study discloses new antiviral components of BM and suggests a passive transfer of these protective factors to the infant via breastfeeding, especially in the first few days of lactation.

Ginsenoside-Rb2 and 20(S)-Ginsenoside-Rg3 from Korean Red Ginseng Prevent Rotavirus Infection in Newborn Mice.

It is well known that Korean red ginseng has various biological activities. However, there is little knowledge about the antiviral activity of Korean red ginseng and its ginsenosides. In this study, we addressed whether oral administration of ginsenoside-Rb2 and -Rg3 is able to protect against rotavirus (RV) infection. The protective effect of ginsenosides against RV infection was examined using an in vivo experiment model in which newborn mice (10-day-old) were inoculated perorally (p.o.) with 1.5 × 106 plaque-forming units/mouse of RV strain SA11. When various dosages of ginsenoside-Rb2 (25-250 mg/kg) were administered 3days, 2 days, or 1 day before virus challenge, treatment with this ginsenoside at the dosage of 75 mg/kg 3days before virus infection most effectively reduced RV-induced diarrhea. In addition, consecutive administration of ginsenoside-Rb2 (75 mg/kg) at 3 days, 2 days, and 1 day before virus infection was more effective than single administration on day -3. The consecutive administration of ginsenoside-Rb2 also reduced virus titers in the bowels of RV-infected mice. In an experiment to compare the protective activity between ginsenoside-Rb2 and its two hydrolytic products (20(S)- and 20(R)-ginsenoside-Rg3), 20(S)-ginsenoside-Rg3, but not 20(R)-ginsenoside-Rg3, prevented RV infection. These results suggest that ginsenoside-Rb2 and its hydrolytic product, 20(S)-ginsenoside-Rg3, are promising candidates as an antiviral agent to protect against RV infection.

Mechanism of Anti-rotavirus Synergistic Activity by Epigallocatechin Gallate and a Proanthocyanidin-Containing Nutraceutical.

Epigallocatechin gallate (EGCG) of green tea and the nutraceutical CystiCran®-40 (containing 40% proanthocyanidins) of the cranberry plant have been associated with antiviral activity. The purpose of this work was to determine the mechanism of antiviral synergy between each compound. Coliphage T4II (phage T4) and the rotavirus strain SA-11(RTV) were used as model virus systems. Individual and combined flavonoids structural and molecular weight analyses were performed by NMR and HPCL/MS, respectively. A suboptimal concentration of EGCG or C-40 alone or in combination reduced phage infectivity by ≤10%. Similarly, EGCG (30 µg/ml) and C-40 (25 µg/ml), respectively, reduced RTV titers by 3 and 13%. However, RTV titers were reduced by 32% (p < .05) with both flavonoids used in combination. RTV was not recognized in host cells by electron microscopy 24-h post-inoculation. NMR and HPLC/MS findings revealed significant structural and potential changes in molecular weight of the flavonoids in complex.

Two new biphenyls from the stems of Garcinia tetralata.

Two new biphenyls (1 and 2) and three known xanthones (3-5) were isolated from the ethanol extract of the stems of Garcinia tetralata. Structural elucidations of 1-2 were elucidated by spectroscopic methods including extensive 1D- and 2D-nuclear magnetic resonance spectroscopy techniques. Compounds 1-2 showed anti-rotavirus activities with SI above 10.

In vitro screening for antiviral activity of Turkish plants revealing methanolic extract of Rindera lanata var. lanata active against human rotavirus.

BACKGROUND: Human rotavirus (HRoV) is the leading cause of severe gastroenteritis in infants and children under the age of five years. No specific antiviral drug is available for HRoV infections and the treatment of viral diarrhea is mainly based on rehydration and zinc treatment. In this study, we explored medicinal plants endemic to Turkey flora as a source of anti-HRoV compunds. METHODS: We performed an antiviral screening on Ballota macrodonta, Salvia cryptantha and Rindera lanata extracts by focus reduction assay. The extract with the highest selectivity index (SI) was selected; its antiviral activity was further confirmed against other HRoV strains and by virus yield reduction assay. The step of viral replicative cycle putatively inhibited was investigated by in vitro assays. RESULTS: The methanolic extract of R. lanata (Boraginaceae) showed the most favourable selectivity index. This extract exhibited a dose-dependent inhibitory activity against three different HRoV strains (EC50 values ranging from 5.8 µg/ml to 25.5 µg/ml), but was inactive or barely active against other RNA viruses, namely human rhinovirus and respiratory syncytial virus. The R. lanata extract targets the early steps of HRoV infection, likely by hampering virus penetration into the cells. CONCLUSION: These results make the R. lanata methanolic extract a promising starting material for a bioguided-fractionation aimed at identifying anti-HRoV compounds. Further work is required to isolate the active principle and assess its clinical potential.

Medicinal plants and natural molecules with in vitro and in vivo activity against rotavirus: A systematic review.

BACKGROUND: Rotaviruses can cause life-threatening health disorders, such as severe dehydrating gastroenteritis and diarrhea in children. Vaccination is the main preventive strategy to reduce rotavirus diarrhea and the severity of episodes, but vaccines are not fully effective and new episodes may occur, even in vaccinated children. The WHO recommends oral rehydration therapy and zinc supplementation for rotavirus-induced diarrhea management. There is little preclinical evidence to support the use of phytotherapeutics in the management of rotaviral infections. PURPOSE: We aim to review the use of medicinal plants and natural molecules in the management of rotavirus infections in experimental studies. METHODS: Articles, published in the English language between 1991 and 2016, were retrieved from PubMed, Scopus and Web of Science using relevant keywords. The scientific literature mainly focusing on plant natural products with therapeutic efficacies against experimental models of rotavirus, were identified and tabulated. In addition, an assessment of the reliability of animal experiments was determined under ``Risk of Bias'' criteria. CHAPTERS: After an initial search and a revision of the inclusion criteria, 41 reports satisfied the objectives of the study. 36 articles were found concerning the anti-rotaviral potential in rotavirus infected cell lines. Among the active secondary metabolites screened for rotavirus inhibition, the polyphenols of flavonoid structure had acquired the highest number of studies in our survey, compared to phenolic acids, stilbenoids, tannins, pectins, terpenoids and flavonoid glycosides. Also, many phytochemicals reduced the efficacy of viral capsid proteins foremost to their elimination and improved the tendency of host-cell inhibiting virus absorption or by prevention of viral replication. Furthermore, five in vivo studies reported that herbs, as well its components, reduced the duration and severity of diarrhea in mice and piglets. The anti-rotavirus efficacy were highlighted based on improvements in reduction on liquid stool, fecal virus shedding, small intestinal histology, levels of inflammation related cytokines and signaling receptors. However, the quality of the experiments in animal studies contained certain types of bias in terms of how they were conducted and reported. CONCLUSION: We identified and summarized studies on medicinal plants and natural molecules having anti-rotavirus activity in order to further future developments of cures for rotavirus gastroenteritis.

Vitamin D3 supplementation alleviates rotavirus infection in pigs and IPEC-J2 cells via regulating the autophagy signaling pathway.

Vitamin D had an anti-infection effect and benefited to the intestinal health. Autophagy signaling pathway was regulated by vitamin D3 to inhibit the infection of human immunodeficiency virus type-1. Rotavirus (RV) was a major cause of the severe diarrheal disease in young children and young animals. Although evidence suggested that vitamin D3 attenuates the negative effects of RV infection via the retinoic acid-inducible gene I signaling pathway, little is known of its antiviral effect whether through the regulation of autophagy. The present study was performed to investigate whether vitamin D3 alleviates RV infection in pig and porcine small intestinal epithelial cell line (IPEC-J2) models via regulating the autophagy signaling pathway. RV administration increased the Beclin 1 mRNA abundance in porcine jejunum and ileum. 5000 IU/kg dietary vitamin D3 supplementation greatly up-regulated LC3-II/LC3-I ratios and PR-39 mRNA expression under the condition of RV challenged. The viability of IPEC-J2 was significantly inhibited by RV infection. Incubation with 25-hydroxyvitamin D3 significantly decreased the concentrations of RV antigen and non-structural protein 4 (NSP4), and up-regulated the mRNA expression of Beclin 1 and PR-39 in the RV-infected IPEC-J2 cells. And then, based on the 25-hydroxyvitamin D3 treatment and RV infection, LC3-II mRNA expression in cells was inhibited by an autophagy inhibitor 3-methyladenine (3-MA). Bafilomycin A1 (Baf A1, a class of inhibitors of membrane ATPases, inhibits maturation of autophagic vacuoles) treatment numerically enhanced the LC3-II mRNA abundance, but had no effect on NSP4 concentration. Furthermore, 25-hydroxyvitamin D3 decreased the p62 mRNA expression and increased porcine cathelicidins (PMAP23, PG1-5 and PR-39) mRNA expression in the RV-infected cells. Taken together, these results indicated that vitamin D3 attenuates RV infection through regulating autophagic maturation and porcine cathelicidin genes expression.

Egg yolk IgY antibodies: A therapeutic intervention against group A rotavirus in calves.

Bovine group A rotavirus (RVA) is considered the major cause of diarrhea in intensively reared neonatal calves. Chicken egg yolk antibodies (IgY) are efficient in protecting neonatal calves from RVA diarrhea; however, the value of this intervention in calves once diarrhea has appeared is unclear. The aim of the present study was to evaluate the application of RVA-specific IgY as a passive treatment in those cases. The experimental groups were: G1=RVA-specific IgY treatment; G2=no Ab treatment; and G3=colostrum deprived+no Ab treatment. IgY treatment significantly reduced virus shedding, diarrhea duration and severity compared to G2 and G3 calves. However, it caused a partial suppression of systemic Ab responses to RVA that could be associated with less severe diarrhea. The oral treatment with IgY for 7days was associated with significantly higher antibody secreting cell responses in the calves compared with other groups of animals.

In vitro antiviral activity against rotavirus and astrovirus infection exerted by substances obtained from Achyrocline bogotensis (Kunth) DC. (Compositae).

BACKGROUND: Achyrocline bogotensis has been traditionally used to treat infections of skin, respiratory, tract urinary and other infections, but not to treat viral gastrointestinal disease. In this study, this Colombian native medicinal plant was investigated by its in vitro anti-rotavirus and anti-astrovirus activity. METHODS: Several extracts and fractions phytochemically obtained from A. bogotensis were evaluated initially for their cell toxicity on MA104 and Caco2 cells and then for their anti-rotavirus (RRV) and anti-astrovirus (Yuc8) activity following three strategies: pre-treatment of cells (blocking effect), direct viral activity (virucidal effect) and post-treatment of infected cells (reduction of viral yield post-infection). In addition qualitative chemical studies were developed for the active compounds. RESULTS: Non-toxic concentrations of a fraction obtained exhibited antiviral activity against both viruses characterized by a virucidal effect and by the reduction of the infectious particles produced post-infection. Steroids, sterols, terpenes, phenols, flavonoids and sesquiterpenlactones were identified qualitatively in the active fraction. CONCLUSIONS: A. bogotensis contains substances with in vitro antiviral activity against rotavirus and astrovirus. This study confirms their anti-microbial properties and describes by the first time its antiviral activity in vitro.

Bovine κ-casein inhibits human rotavirus (HRV) infection via direct binding of glycans to HRV.

Human rotavirus (HRV) is a major etiologic agent of severe infantile gastroenteritis. κ-Casein (κ-CN) from both human and bovine mature milk has been reported to have anti-HRV activity; however, the mechanism of this activity is poorly understood. The present study examined the molecular basis for the protective effect of bovine κ-CN derived from late colostrum (6-7 d after parturition) and from mature milk. Among the components of casein, κ-CN is the only glycosylated protein that has been identified. Therefore, we investigated whether the glycan residues in κ-CN were involved in the anti-HRV activity. Desialylated CN obtained by neuraminidase treatment exhibited anti-HRV activity, whereas deglycosylated CN obtained by o-glycosidase treatment lacked antiviral activity, indicating that glycans were responsible for the antiviral activity of CN. Furthermore, an evanescent-field fluorescence-assisted assay showed that HRV particles directly bound to heated casein (at 95°C for 30 min) in a viral titer-dependent manner. Although the heated κ-CN retained inhibitory activity in a neutralization assay, the activity was weaker than that observed before heat treatment. Our findings indicate that the inhibitory mechanism of bovine κ-CN against HRV involves direct binding to viral particles via glycan residues. In addition, heat-labile structures in κ-CN may play an important role in maintenance of κ-CN binding to HRV.

Synthesis, pharmacological activity evaluation and molecular modeling of new polynuclear heterocyclic compounds containing benzimidazole derivatives.

Novel heterocyclic compounds containing benzimidazole derivatives were synthesized from 2-(1H-benzimidazol-2-yl) acetonitrile (1) and arylhydrazononitrile derivative 2 was obtained via coupling of 1 with 4-methyl phenyldiazonium salt, which was then reacted with hydroxylamine hydrochloride to give amidooxime derivative 3. This product was cyclized into the corresponding oxadiazole derivative 4 upon reflux in acetic anhydride. Compound 4 was refluxed in DMF in the presence of triethylamine to give the corresponding 5-(1H-benzimidazol-2-yl)-2-p-tolyl-2H-1,2,3-triazol-4-amine 6. Treatment of compound 6 with ethyl chloroformate afforded 2,6-dihydro-2-(4-methylphenyl)-1,2,3-triazolo[4",5"-4',5']pyrimido[1,6-a]benzimidazole-5(4H)-one (8). 1,2-bis(2-cyanomethyl-1H-benzimidazol-1-yl)ethane-1,2-dione (10) was synthesized via the condensation reaction of 2-(1H-benzimidazol-2-yl) acetonitrile (1) and diethyloxalate. The reactivity of compound 10 towards some diamine reagents was studied. The in vitro antimicrobial activity of the synthesized compounds was investigated against several pathogenic bacterial strains such as Escherichia coli O157, Salmonella typhimurium, E. coli O119, S. paratyphi, Pseudomonas aeruginosa, Staphylococcus aureus, Listeria monocytogenes and Bacillus cereus. The results of MIC revealed that compounds 12a-c showed the most effective antimicrobial activity against tested strains. On the other hand, compounds 12a, b exhibited high activity against rotavirus Wa strain while compounds 12b, c exhibited high activity against adenovirus type 7. In silico target prediction, docking and validation of the compounds 12a-c were performed. The dialkylglycine decarboxylase bacterial enzyme was predicted as a potential bacterial target receptor using pharmacophore-based correspondence with previous leads; giving the highest normalized scores and a high correlation docking score with mean inhibition concentrations. A novel binding mechanism was predicted after docking using the MOE software and its validation.

An evaluation of the inhibitory effects against rotavirus infection of edible plant extracts.

BACKGROUND: Rotaviruses are the single most important cause of severe diarrhea in young children worldwide. The developments of specific, potent and accessible antiviral treatments that restrain rotavirus infection remain important to control rotavirus disease. METHODS: 150 plant extracts with nutritional applications were screened in vitro on MA-104 cells for their antiviral activity against rhesus rotavirus (RRV). One extract (Aspalathus linearis (Burm.f.) R.Dahlgren) was also tested for its effect on the loss of transepithelial resistance (TER) of Caco-2 cells caused by simian rotavirus (SA-11) infection. RESULTS: Aqueous extracts of Nelumbo nucifera Gaertn. fruit, Urtica dioica L. root, Aspalathus linearis (Burm.f.) R.Dahlgren leaves, Glycyrrhiza glabra L. root and Olea europaea L. leaves were found to have strong significant antiviral activity with a 50% inhibitory concentration (IC50) < 300 µg/ml. The pure compound 18ß-glycyrrhetinic acid from Glycyrrhiza glabra was found to have the strongest antiviral activity (IC50 46 µM), followed by luteolin and vitexin from Aspalathus linearis (IC50 respectively 116 µM and 129 µM) and apigenin-7-O-glucoside from Melissa officinalis (IC50 150 µM). A combination of Glycyrrhiza glabra L. + Nelumbo nucifera Gaertn. and Urtica dioica L. + Nelumbo nucifera Gaertn. showed synergy in their anti-viral activities. Aspalathus linearis (Burm.f.) R.Dahlgren showed no positive effect on the maintenance of the TER. CONCLUSIONS: These results indicate that nutritional intervention with extracts of Nelumbo nucifera Gaertn., Aspalathus linearis (Burm.f.) R.Dahlgren, Urtica dioica L., Glycyrrhiza glabra L. and Olea europaea L. might be useful in the treatment of diarrhea caused by rotavirus infection.

Inhibition of rotavirus infection in cultured cells by N-acetyl-cysteine, PPARγ agonists and NSAIDs.

Although the current rotavirus vaccines have shown good tolerance and significant efficacy, it would be useful to develop alternative or complementary strategies aimed at preventing or treating acute diarrhoeal disease caused by this viral agent. A variety of antiviral strategies other than vaccines have been assayed for rotavirus infection management. The recently demonstrated sensitivity of rotavirus infectivity to thiol/disulfide reagents prompted assays for screening drugs that potentially affect cellular redox reactions. MA104 or Caco-2 cells were inoculated with the rotavirus strains RRV, Wa, Wi or M69 and then incubated with different concentrations of drugs belonging to a selected group of 60 drugs that are currently used in humans for purposes other than rotavirus infection treatment. Eighteen of these drugs were able to inhibit rotavirus infectivity to different extents. A more systematic evaluation was performed with drugs that could be used in children such as N-acetylcysteine and ascorbic acid, in addition to ibuprofen, pioglitazone and rosiglitazone, all of which affecting cellular pathways potentially needed by the rotavirus infection process. Evidence is provided here that rotavirus infectivity is significantly inhibited by NAC in different cell-culture systems. These findings suggest that NAC has the potential to be used as a therapeutic tool for treatment and prevention of rotavirus disease in children.

Screening of Brazilian medicinal plants for antiviral activity against rotavirus.

ETHNOPHARMACOLOGICAL RELEVANCE: Brazilian medicinal plants traditionally used for the treatment of diarrhoea were investigated for their in vitro antiviral activity against the simian rotavirus SA11. MATERIALS AND METHODS: The ethanolic crude extracts of plants collected in the cerrado of Minas Gerais, Brazil were submitted to phytochemical screening. The cytotoxicity of the extracts was inferred by cellular morphologic alterations. Antiviral activity was assessed by the ability of the extracts to inhibit the cytopathic effect (CPE) of rotavirus on the treated cells. RT-PCR was performed to confirm and/or confront antiviral assay data. RESULTS: The maximum non-toxic concentration ranged from 50 to 500 µg/mL. All extracts were toxic at a concentration of 5000 µg/mL but no extract showed cytotoxicity at 50 µg/mL. The species Byrsonima verbascifolia, Myracrodruon urundeuva, Eugenia dysenterica and Hymenaea courbaril exhibited the strongest in vitro activity against rotavirus. Their extracts prevented the formation of CPE, and RT-PCR analysis detected no amplification of genetic material from rotavirus. Tannins, flavonoids, saponins, coumarins and terpenes were the major classes of natural products found in the leaf extracts that showed antiviral activity. CONCLUSION: Among the species studied, Byrsonima verbascifolia, Eugenia dysenterica, Hymenaea courbaril and Myracrodruon urundeuva showed potential activity against rotavirus and are worthy of further study. The present study corroborates ethnopharmacological data as a valuable source in the selection of plants with antiviral activity and to some extent validates their traditional uses.